Glycosphingolipids (GSL's) are ubiquitous membrane components found in all animal cells and most plant cells, including yeasts. GSL's play crucial functional roles in cell-cell recognition, cell adhesion, and modulation of transmembrane signaling [1]. Patterns of GSL expression change dramatically during the processes of ontogenesis, oncogenesis and differentiation of stem cells into phenotypically terminal-stage cells. The knowledge of GSL distribution and expression was enriched greatly by introduction of the monoclonal antibody approach and many developmentally-regulated, tumor-associated GSL antigens have been identified on a chemical basis [2,3].
Apoptosis (programmed cell death) is a fundamental energy-dependent process in all multicellular organisms. Apoptosis helps control tissue size, shape and balance of cell number during morphogenesis, and plays an important role in oncogenic progression [4,5] and in other pathological processes such as reperfusion injury [6] and glomerulonephritis [7]. Aging neutrophils also have been shown to undergo apoptosis spontaneously [8,9]. Apoptotic senescent neutrophils are recognized and phagocytized by macrophages. The process has been suggested to represent a mechanism in vivo to limit neutrophil-mediated tissue injury at inflamed sites.
Various cytokines released during inflammation regulate the survival of neutrophils at the lesion either by promoting or by inhibiting cell death. Inflammatory mediators, such as endotoxic lipopolysaccharide, complement factor 5a and granulocyte-macrophage colony-stimulating factor, markedly inhibited neutrophil apoptosis [10]. In contrast, TNF-.alpha., a potent neutrophil activator [11-13], has been shown to accelerate the rate of neutrophil apoptosis [14]. TNF-e responsive sphingomyelin hydrolysis and ceramide generation have been reported to be implicated in a signal transduction pathway that mediates induction of apoptosis by TNF-.alpha. in U-937 cells [15,16].
Calcium/phospholipid-dependent protein kinase (protein kinase C; PKC) is involved in intracellular signaling processing including those of cellular proliferation and differentiation in a variety of cells [17]. In hematopoietic cell systems, treatment with pharmacologic inhibitors of PKC causes the growth inhibition of both normal [18] and leukemic [19] progenitors. Several investigators reported that exposure to PKC inhibitors, such as H7 and staurosporine, induced apoptosis in HL-60 promyelocytic leukemia cells [20,21], MOLT-4 lymphoid leukemia cells and normal lymphocytes [22] and a variety of neoplastic cell lines [23]. In addition, activation of PKC by exposure to PMA prevented growth factor-deprived hematopoietic cells from undergoing apoptotic cell death [24]. Those observations suggest the potential role of PKC in the regulation of apoptosis.
Sphingosine, a sphingolipid breakdown product, has been shown to inhibit PKC in vitro and in cells [25-27]. Sphingosine and a catabolite thereof, DMS, had inhibitory effects on in vitro as well as in vitro tumor cell growth [28-30]. DMS also has an inhibitory effect on PKC activity as well as sphingosine [31].
Sphingosine and DMS appear to inhibit cell growth and exert a cytotoxic activity. It has been postulated that sphingosine functions as an endogenous modulator of PKC and plays important roles in cell growth, differentiation and oncogenesis [27,32]. However, because of difficulty of measuring changes in cellular sphingosine levels in response to biological stimuli, whether sphingosine functions physiologically in mediating biologic processes including growth suppression has not been determined [33].
On the other hand, human myeloid leukemia cell lines, including HL-60 cells, have retained the capacity to respond to inducers of differentiation with the cessation of growth and appearance of a more mature phenotype [34]. Recently, evidence of apoptosis has been described in HL-60 cells during both PMA-induced macrophage differentiation [35] and retinoic acid-induced neutrophilic differentiation [36].
However, it remains unclear what initiates apoptosis during cell differentiation. It is of interest to determine whether sphingosine, a potent endogenous PKC inhibitor, is involved in the mechanism of loss of proliferative capacity and induction of apoptosis during differentiation, since pharmacologic PKC inhibitors cause inhibition of cell growth and apoptosis.